Unitary pharmaceutical dosage form

ABSTRACT

In accordance with this invention a novel pharmaceutical product containing efavirenz, emtricitabine and tenofovir DF are provided as a multicomponent unitary oral dosage form, component 1 comprising tenofovir DF (and, optionally, emtricitabine) and component 2 comprising efavirenz, wherein components 1 and 2 are in a stabilizing configuration. In preferred embodiments component 1 is made by dry granulation.

BACKGROUND OF THE INVENTION

This application relates to products for the treatment of viral infections, in particular HIV infections, using the known antiviral compounds efavirenz (tradename Sustiva, also known as EFV), emtricitabine (tradename Emtriva, also known as FTC) and tenofovir DF (disoproxil fumarate, also known as TDF) (tradename Viread, sold in combination with emtricitabine under the tradename Truvada).

The Truvada product is produced by wet granulation of emtricitabine and tenofovir DF (WO 04/64845), which under the circumstances produces a chemically stable dosage form. This product does not contain efavirenz.

HIV therapy using efavirenz as well as emtricitabine and tenofovir DF has been considered desirable (hereafter “triple combination”; see WO 04/64845). Manufacturing a commercially viable triple combination product, however, would require that the final product meet stringent FDA requirements for bioequivalence to the commercial products, Viread (tenofovir disoproxil fumarate), Emtriva (emtricitabine), and Sustiva (efavirenz), and that the tablet be of suitable size for patients to easily swallow.

Initial efforts to simply combine the three drugs (active pharmaceutical intermediates, or APIs) into a unitary, essentially homogeneous composition manufactured by wet granulation failed to produce a chemically stable tablet. The tenofovir DF in this combination tablet was highly unstable and rapidly degraded in stability studies. The efavirenz formulation was unexpectedly incompatible with tenofovir DF, a result now attributed to the surfactant (sodium lauryl sulfate) found in the efavirenz portion of the formulation.

Another attempt was made to produce the triple combination, this time using a dry granulation of the three part combination and omitting the surfactant. This resulted in a tablet that failed to achieve bioequivalence with respect to efavirenz in human clinical trials. The peak efavirenz concentration in the blood stream and total drug exposure (Cmax and AUC) were both below the parameters determined for the commercial comparator, Sustiva (efavirenz) tablets. The inventors concluded that at least the surfactant in the triple combination (efavirenz/emtricitabine/tenofovir disoproxil fumarate) tablets was necessary to achieve bioequivalence to Sustiva.

Next, combination tablets were manufactured by wet granulating the efavirenz component with the surfactant and other excipients, separately manufacturing the Truvada component using dry granulation, mixing the granulates together, compressing the mixture into tablets, and then film-coating the tablets. Unexpectedly, this approach also failed to produce the desired bioequivalence in between the commercial product, Sustiva (efavirenz), and clinical trial material (i.e., proposed commercial triple combination product). A novel and inventive step was needed to overcome the shortcomings of more straight-forward approaches to a triple combination dosage form.

Copending U.S. Ser. No. 60/771,353 (filed of even date and expressly incorporated herein by reference) is directed to solving another obstacle encountered in the preparation of the triple combination dosage form, that of reducing the size of the combined product. While the prior art reports the successful manufacture of chemically stable Truvada preparations (WO04/64845), these preparations contain relatively low proportions of excipient to API. Increasing the proportion of excipients and wet granulating the three API combination unexpectedly resulted in a preparation in which the tenofovir DF was highly unstable. As reported in U.S. Ser. No. 60/771,353, it was believed that use of sufficient water to accomplish the wet granulation of efavirenz (which has relatively low solubility in comparison to emtricitabine and tenofovir DF) caused the latter two APIs to dissolve into a eutectic mixture. The eutectic mixture dried during granulation to form a glassy or amorphous product in which the tenofovir DF is chemically unstable in comparison to the crystalline API. Supplying enough excipient to ameliorate the effect of the excess water was not consistent with the objective of obtaining a triple combination oral dosage form of manageable proportions.

As described further in U.S. Ser. No. 60/771,353, this obstacle was overcome by dry granulating the emtricitabine and tenofovir DF composition, i.e., granulating the composition without contacting same with a destabilizing amount of liquid water. Omitting water (particularly, liquid water) or reducing the presence of water to an insubstantial amount eliminates the disadvantages formation of a eutectic mixture and enhances the stability of the resulting pharmaceutical product.

Despite the advantages conferred by dry granulation of the emtricitabine/tenofovir DF component, it was still necessary to overcome the unexpected incompatibility of tenofovir DF and the surfactant used in the Sustiva formulation.

SUMMARY OF THE INVENTION

In accordance with this invention, the stability and bioequivalence objective for the triple combination product have been achieved by providing a multicomponent dosage form, one component comprising tenofovir DF and, optionally, emtricitabine, and the other comprising at least efavirenz. Another embodiment of the invention is a dosage form comprising a tenofovir DF component and a surfactant component not in destabilizing contact with the tenofovir DF component.

DETAILED DESCRIPTION OF THE INVENTION

The dosage form of this invention comprises efavirenz, emtricitabine and tenofovir DF. As noted, tenofovir DF and efavirenz are in separate components, Emtricitabine generally is included in the tenofovir DF component, but in other embodiments the emtricitabine is present in its own component, or is mixed with the efavirenz component. Its disposition is not critical to the practice of this invention. All that is necessary is that emtricitabine be present in the dosage form and that the tenofovir DF component be substantially separated from the surfactant in the efavirenz component. Any method, additive, process feature or configuration that suitably minimizes the contact of surfactant with tenofovir DF is suitable in the practice of this invention.

The term “component” means a physically discrete unit or compartment which is associated physically with and in contact with other components. This does not mean that the units or compartments are physically not in contact. In fact, it generally is preferred that they are in physical contact and form a unitary device, article or composition. The degree of association is only that which is needed to facilitate the oral consumption of the composition as a single dosage form. This invention does not include, for example, patient packs with the Sustiva and Truvada products in separate wells or containers, or other associations which are essentially packaging solutions alone (although, of course, the compositions at this invention optionally are packed or packaged in any conventional fashion suitable under the circumstances).

Typically, the components of the dosage form of this invention conveniently are organized in multiple layers, ordinarily a bilayer as shown in the exemplified embodiment. However, if emtricitabine is present in its own component then the dosage form will constitute at least a trilayer structure. There need not be a single component for each drug (for example, the dosage forms optionally include 2 layers for each of the components, for a total of 6). Thus, the dosage unit includes laminates of many components. There do not need to be equal numbers of each component, e.g., layers, for each drug or drug combination so long as the total dosage of all components in sum is the desired amount.

Other means for spatially organizing components are suitable so long as the desired degree of separation of tenofovir DF and surfactant is accomplished. For example, rather than forming planar layers along the axis of a tablet, the components optionally are organized in an annular fashion, with each ring or cylinder containing a separate component. Another alternative is to employ a press coating process to associate the components.

The components generally are in direct contact with one another, i.e., no barrier or protective layer is present between them. In other embodiments, a barrier is introduced between the incompatible components. A suitable example of this embodiment of the invention would be a multi-compartment capsule in which the incompatible components are distributed into separate compartments. Alternatively, a tablet is optionally provided that contains one encapsulated component disbursed or distributed within the incompatible component. In general, intimate, direct admixture of the incompatible components is undesirable unless means are provided to protect the tenofovir DF component from surfactant.

In typical embodiments the components of the dosage form of this invention are spatially organized so as to not place the tenofovir DF component into destabilizing contact with the surfactant in the efavirenz component. “Destabilizing” means any contact between tenofovir DF and the surfactant that is capable causing pharmaceutically unacceptable degradation of tenofovir DF. A stabilizing configuration is any spatial organization of the tenofovir DF and efavirenz components that does not result in the generation of a “pharmaceutically unacceptable amount” of any one of the following degradation products. A destabilizing contact is a spatial organization that results in the generation of any of the following degradation products in a “pharmaceutically unacceptable amount”.

The spatial geometry and conditions of the permitted contact between tenofovir DF and surfactant-containing component are essentially unlimited. This spatial geometry is termed a “stabilizing configuration” or, stated differently, is a configuration that does not contain a “destabilizing contact” as defined below. There are many ways in which the central observation of this invention (that is, that sodium lauryl sulfate destabilizes tenofovir DF) can be harnessed to prevent the generation of pharmaceutically unacceptable levels of degradation of tenofovir DF.

In addition, when emtricitabine is present in the tenofovir DF component, the permitted contact also that which does not produce pharmaceutically unacceptable amounts of emtricitabine degradation product.

“Degradation” of tenofovir DF is the generation—in pharmaceutically unacceptable amounts—of at least one of the degradation products mono-POC PMPA, dimer or mixed dimer. “Degradation” of FTC is defined as the generation—in pharmaceutically unacceptable amounts, of FTU. These degradation products are shown below.

Mono-POC PMPA

Dimeric Degradation Products

Dimer

Mixed Dimmer

FTU has the structure

A “pharmaceutically unacceptable amount” is defined as the following amounts of each degradation product. Degradation products optionally are assayed in either an absolute or incremental amount. This absolute or total amount of degradation product is simple the amount found in the test article. The incremental amount is the additional amount of degradation product appearing in the product over that which was present (if any) in the API starting material. Moreover, the amount of degradation product optionally is measured at two points in time. One is at the time of release into the marketplace. The other is after exposure to storage conditions under the conditions described below, i.e., the shelf life as set forth below.

Total Amounts at Release (First Commercial Sale)

No more than about 3%, ordinarily about 1.5%, of mono-POC PMPA.

No more than about 1%, ordinarily about 0.5of Dimer.

No more than about 0.5%, ordinarily about 0.25% of Mixed Dimer.

Less than about 0.5%, ordinarily about 0.2% of FTU.

Total Amounts at Shelf Life (Storage at 25° C./60% RH for 24 mo.)

No more than about 10%, ordinarily about 5% of mono-POC PMPA.

No more than about 2%, ordinarily about 1% of Dimer.

No more than about 2%, ordinarily about 1% of Mixed Dimer.

No more than about 4%, ordinarily about 2% of FTU.

Incremental Amounts at Release (First Commercial Sale)

No more than about 2%, ordinarily about 0.5%, of mono-POC PMPA.

No more than about 0.6%, ordinarily about 0.1% of Dimer.

No more than about 0.3%, ordinarily about 0.05% of Mixed Dimer.

Less than about 0.4%, ordinarily about 0.1% of FTU

Incremental Amounts at Shelf Life (Storage at 25° C./60% RH for 24 mo.)

No more than about 9%, ordinarily about 4% of mono-POC PMPA.

No more than about 1.6%. ordinarily about 0.6% of Dimer.

No more than about 1.8%, ordinarily about 0.8% of Mixed Dimer.

No more than about 3.9%, ordinarily about 1.9% of FTU.

The percentage of degradation products is the amount of degradation product as measured by HPLC retention time comparison. In the HPLC retention time comparison, the retention time of the main peaks observed in the tablets is required to be within 2% of the retention time of the main peaks in the a reference standard preparation containing efavirenz, emtricitabine, and tenofovir DF in an assay which has been shown to be specific for efavirenz, emtricitabine, and tenofovir DF. The percentage is determined be dividing the total amount of tenofovir DF plus the three degradation products into the amount of individual degradation product as determined by the HPLC assay.

These parameters are employed to evaluate whether a test composition has met the requirements of a stabilizing contact. For example, a triple combination dosage form optionally is designed as a shaped article comprising slugs of compressed granules of the tenofovir DF component dispensed within a matrix of the efavirenz component. A variety of slug sizes might be used in making the composition. This constellation of potential products then would be tested, or stored under the conditions above and then tested, to assay the generation of tenofovir DF and/or FTC degradation products. If the resulting product upon release did not contain more than the specified approximate limits of any one or more of the 4 containments listed under any of the 4 assay paradigms above, then the contact would be considered stabilizing. Of course, the artisan may adopt more stringent standards, but this will be a matter of choice and shall not limit the scope of this invention.

In preferred embodiments the emtricitabine and tenofovir DF are combined and this component is prepared by dry granulation (U.S. Ser. No. 60/771,353). In preferred embodiments, a composition comprising dry granulated tenofovir DF and emtricitabine is employed in one component of the dosage forms of this invention.

Dry granulation is a well-known pharmaceutical manufacturing process per se. In general, API is combined with excipients and lubricant excipient and then compressed to form a mass. This mass typically is then comminuted or milled, then sieved to obtain the desired size of particle. The granular product is compressed into tablets, filled into capsules or otherwise formed into a unitary dosage form in conventional fashion.

Compression into a mass is accomplished by conventional equipment. Typically, the API and excipients are passed through a roller apparatus for compaction. However, other means for compacting the API mixture, e.g., compaction into slugs (or “slugging”), optionally are used.

A dry granulation process is one in which a dry composition of the API and selected excipient(s) is compressed to form a mass, which is comminuted or milled if necessary, and then optionally sieved to produce the desired size granules. Compression into a mass is accomplished by conventional equipment. Typically, the API and excipients are passed through a roller apparatus for compaction. However, other means for compacting the API mixture, e.g., compaction into slugs (or “slugging”), can be used.

A composition comprising dry granulated emtricitabine and tenofovir DF is the product of a dry granulation process. This composition essentially retains the crystalline APIs and is substantially free of dried eutectic emtricitabine/tenofovir DF. It typically will contain less than about 15% by weight dried eutectic mixture, ordinarily less than about 10% and generally less than about 5%.

The dry granulation process is conducted in the absence of a destabilizing amount of water, “destabilizing” being that amount of liquid water that is capable causing pharmaceutically unacceptable degradation of tenofovir DF and/or FTC as defined herein. If the dosage form of this invention includes a dry granulated emtricitabine/tenofovir DF component, then the amount of permitted degradation product in the final dosage form is still the same as that which is set forth above, i.e., the amount of water exposure and contact, together or alone, are not to result in degradation products failing to meet the standards described above. It is an option, of course, to test the dry granulates for their level of degradation product first, and if they pass, then to formulate them into the dosage form of this invention and then determine if the contact results in any increase in degradation products that takes the resulting dosage form outside the parameters established.

Bound, entrained or absorbed water are commonly present in excipients. This water will not significantly adversely affect the stability of tenofovir DF and thus is not excluded from the dry granulates optionally used in the dosage form of this invention. In general, liquid water (added or generated in situ) from any source, e.g., chemical reactions, condensation, entrained ice, or the like is to be excluded from the granulation. However, minor amounts of liquid water optionally are added during granulation. These amounts typically would be less than about 5% by weight, ordinarily less than about 1% by weight, however the water is generated or supplied. Water is present in the final granulation product up to about 10% by weight (Karl Fischer), but preferably is less, as low as 0.1% by weight. However, permitted quantities of water may vary depending upon other factors in the granulation, e.g., excipient type, temperature and so forth. For example, if a hygroscopic excipient is included this will convert added water into a bound form. All that is necessary is that the water not result in degradation of tenofovir DF and/or emtricitabine in the final product. In general, water is excluded both from the pregranulation stage (preparation of the composition to be used directly in the granulation) as well as during the granulation process itself.

Absence of water or “dry” does not mean the absence of liquid. Granulations with organic solvents are also feasible provided that destabilizing amounts of water are excluded.

Dry granulation results in a product that contains minimal amounts of water. The amount of water in the product granulate or dosage forms made therefrom are measured by loss on drying (LOD) or by the Karl Fischer method. The LOD of compositions of this invention are about 15%, about 10%, about 5% or typically less than about 3% by weight. The Karl Fischer water is about from 0.1 to 10% by weight, usually less than about 5% by weight, or less than about 2%. The amount of water in the final preparations, as opposed to the granulates, is a function of granulate water as well as minor amounts of water used during subsequent process steps such as coating. These amounts of water added in later steps than granulation generally will not affect the stability of the emtricitabine/tenofovir DF APIs, and therefore are subject to considerable permitted variation.

The manufacturing process described below is directed to the preparation of a triple combination tablet containing efavirenz, emtricitabine and tenofovir DF. In this particular embodiment the last two drugs are emplaced in a portion of the tablet which is separate from, but in contact with, the portion of the tablet containing efavirenz. It will be understood, however, that the emtricitabine and tenofovir DF component of the tablet, which is an embodiment of this invention, optionally is manufactured as a stand-alone product and not necessarily in assembly with an efavirenz component. In this option, the emtricitabine/tenofovir DF dry granulation intermediate described below is simple compressed into tablets or conventionally processed into other conventional unitary dosage forms such as capsules, cachets, suppositories, or the like.

The dosage forms of this invention are stored in containers, preferably under desiccant such as silica gel in amounts generally sufficient to maintain the RH over the dosage forms at under about 10%, preferably under about 5%.

Materials

The quantitative compositions of the efavirenz powder blend, FTC/TDF powder blend, and film-coated bi-layer EFV/FTC/TDF tablets are listed in Table 1, Table 2, and Table 3. respectively. The quantities of efavirenz, emtricitabine, and tenofovir DF were adjusted for drug content factors (DCF) if the value was less than 0.99 with a concomitant reduction to the quantity of microcrystalline cellulose in each granulation.

TABLE 1 Quantitative composition of efavirenz powder blend % w/w Unit Formula Ingredient of Total (mg/tablet) Efavirenz 38.71 600.0 Microcrystalline Cellulose, NF/EP 11.52 178.6 Hydroxypropyl cellulose, NF/EP 2.48 38.4 Sodium Lauryl Sulfate, USP/EP 0.77 12.0 Croscarmellose Sodium, NF/EP 3.87 48.0 Magnesium Stearate, NF/EP 0.58 9.6 Total for Tablet Core 57.94 898.0

TABLE 2 Quantitative composition of FTC/TDF powder blend % w/w Unit Formula Ingredient of Total (mg/tablet) Emtricitabine 12.90 200.0 Tenofovir Disoproxil Fumarate 19.35 300.0 Microcrystalline Cellulose, NF/EP 5.77 89.5 Croscarmellose Sodium, NF/EP^(a) 3.10 48.0 Magnesium Stearate, NF/EP^(a) 0.94 14.5 Total for Tablet Core 42.06 652.0 ^(a)To be incorporated into both the intragranular and extragranular portions of the formulation during the manufacturing process.

TABLE 3 Quantitative composition of film-coated bi-layer EFV/FTC/TDF Tablets % w/w Unit Formula Ingredient of Total (mg/tablet) Efavirenz Powder Blend 57.94 898.0 FTC/TDF Powder Blend 42.06 652.0 Total for Tablet Cores 100.00 1550.0 Opadry II Pink 3.00 46.5 Purified Water, USP/EP^(a) Total for Film-Coated Tablets 1596.5 ^(a)Water removed during film-coating process.

The excipients were all compendial grade materials.

Efavirenz Wet Granulation

Efavirenz was wet granulated using a Niro-Fielder PMA-400 equipment train. Efavirenz, microcrystalline cellulose and sodium lauryl sulfate (Table 1) were added to the PMA-400 and blended for 3 minutes. Croscarmellose sodium and hydroxyl propyl cellulose (Table 1) were added to the pre-mix and blended for an additional 2 minutes. Purified water was added to form a suitable granulation followed by additional wet massing after water addition. Table 4 lists the summary of granulation parameters used for two representative lots and sub parts. All sub parts used a water to efavirenz ratio of 1.30 except for AB509 Mix C which used a 1.25 ratio of water to efavirenz.

TABLE 4 Efavirenz wet granulation process parameter summary AB507 AB509 Mix Mix Mix Mix Mix Mix Process Parameter A B C A B C Granula- Total Water 33.57 33.56 33.56 33.56 33.56 32.18 tion Added (kg) Ratio of 1.30 1.30 1.30 1.30 1.30 1.25 Water:EFV Total Addition 9:36 9:29 9:24 9:17 9:32 9:02 Time (Min:Sec) Final Impeller 10.4 9.8 8.5 11.3 11.3 9.9 Power (% Load) Wet Total Time 4:00 3:00 3:00 2:00 1:15 2:00 Massing (Min:Sec) Final Impeller 11.6 12.0 11.7 18.0 17.7 10.5 Power (% Load) Drying^(a) Inlet Temperature 70 70 (° C.) Time (Hr:Min) 1:45 1:51 Final Outlet 50 50 Temp. (° C.) Final LOD 0.3 0.8 (%) ^(a)Mixes A, B, and C for each lot were combined before drying.

In general, the wet granules were milled, then dried to an LOD less than or equal to 1.5%. The dried granules were milled and blended with magnesium stearate (Table 1).

The bulk density, particle size, and moisture content by LOD of the efavirenz granulations are listed in the first three lines of Table 5 (the B lot numbers are efavirenz products, the C lot numbers are emtricitabine/tenofovir DF). Particle size was determined by sitting 10-gram samples through 3-inch diameter screens using a sonic sifter (Model L3P, ATM Corporation, Milwaukee, Wis., USA). This following US Standard Mesh sizes (openings) were used: #20 (850 μm), #30 (600 μm), #40 (425 μm), #60 (250 μm), #80 (180 μm), and #250 (63 μm). The agitation and pulse were set at 7 and the sifting time was 5 minutes. The amount of powder retained on the sieves and the fines collector was determined by calculating the difference in weight before and after sifting. The geometric mean particle size was calculated by logarithmic weighting of the sieved distribution.

Bulk density was determined by filling a 100-mL graduated cylinder with sample and calculating the difference in weight between the empty and full graduated cylinder per unit volume.

Moisture content measurements by loss on drying (LOD) were performed by heating a 2.5 g sample at 85° C. for 15 minutes using a heat lamp/balance system (Model LP16/PM400, Mettler-Toledo, Columbus, Ohio, USA).

The granulations had similar bulk densities (0.54 to 0.56 g/mL) and similar geometric mean particle size distributions (215 to 268 μm). The LOD value of the final blend were consistent from 0.98 to 1.80%. The individual sieve distributions for the efavirenz granulations are listed in Table 6.

TABLE 5 Summary of efavirenz powder blend and emtricitabine/tenofovir DF powder blend physical properties Geometric Mean Diameter Gilead Particle Size Bulk Density LOD Lot Number (μm) (g/mL) (%) AB507 247 0.56 1.80 AB508 215 0.55 1.08 AB509 268 0.54 0.98 AC507 330 0.60 0.91 AC508 344 0.60 1.02 AC509 343 0.59 0.99

TABLE 6 Particle size distribution for efavirenz and FTC/TDF powder blends % Weight Retained on Screen^(a) US Standard Screen Size (mesh opening) Gilead 20 30 40 60 80 230 pan Lot (>850 (600 (425 (250 (180 (63 (<63 Number μm) μm) μm) μm) μm) μm) μm) AB507 5.9 10.9 16.2 22.2 11.4 22.6 10.9 AB508 6.1 10.4 15.8 20.0 9.0 20.8 17.9 AB509 9.6 13.3 17.4 20.1 8.9 17.2 13.3 AC507 22.0 19.8 15.2 11.2 4.6 10.5 16.6 AC508 22.1 20.1 15.4 11.6 5.1 10.6 14.9 AC509 22.4 19.7 15.3 11.7 4.8 11.1 14.8

Emtricitabine/Tenofovir DF Dry Granulation

Emtricitabine, microcrystalline cellulose, tenofovir DF, and croscarmellose (Table 2) were blended in a 650 L tote bin using a Gallay blender for 10 minutes. Magnesium stearate (Table 2) was added and blended for an additional 5 minutes. This pre-blend was then transferred to a 320-L Matcon bin fitted with a cone valve discharging station to assist with material transfer into the roller compactor hopper.

The pre-blend was roller compacted using a Gerteis Macro-Pactor model 250/25/3 with 250 mm diameter by 50 mm wide smooth rolls. The roll gap thickness (2 mm), roll spread (10 rpm), compaction force (4 kN/cm), oscillating mill speed (75 rpm clockwise and counterclockwise), and oscillating mill screen opening (1.25 mm) were kept constant for all batches. The oscillating mill angle of rotation was also the same for all lots at 150° clockwise and 140° counterclockwise.

There was no material handling issues among all three batches while feeding into the roller compactor. The entire roller compaction process proceeded without any apparent sign of heat accumulation of the equipment, product build-up, or melting. The granulations then were blended with extragranular croscarmellose sodium (34% of total amount) and magnesium stearate (47% of total amount).

The particle size, bulk density, and LOD of the emtricitabine/tenofovir DF dry granulations were all similar for the three batches and are listed in Table 5 (bottom 3 compartments). The geometric particle sizes were very similar at from 330 to 344 μm. Bulk densities ranged from 0.59 to 0.60 g/mL. The final blend LOD values were consistent from 0.91 to 1.02%. The final powder blends have remarkably consistent physical properties.

The efavirenz and tenofovir DF granulations each have geometric mean particle sizes that optionally range from 100 to 600 μm, bulk densities optionally ranging about from 0.1 to 1 g/mL and LOD values optionally ranging about from 0.1 to 10% by weight.

Final Blends

The mass of efavirenz granulation and extragranular magnesium stearate were adjusted appropriately based on the yield of emtricitabine/tenofovir DF dry granulation. Efavirenz granulation and emtricitabine/tenofovir DF dry granulation were blended in a 3 cubic foot V-blender for 10 minutes. Magnesium stearate was added and blended an additional 5 minutes. Samples of the final powder blend were taken from 10 different locations after blending and analyzed for blend uniformity. The efavirenz and emtricitabine/tenofovir DF final powder blends showed acceptable blend uniformity and homogeneity for all three active ingredients indicating the robustness of the formulation regardless of the particle size or bulk density of emtricitabine/tenofovir DF dry granulations and efavirenz granulations. The granulations and blending procedure would be satisfactory for the formulation of a larger scale.

Tablet Core Compression

Efavirenz/emtricitabine/tenofovir DF final powder blend was compressed into table cores using a Stokes Genesis Model 757, 41 station bilayer tablet press equipped plain-faced upper/embossed “123” lower, capsule-shaped (20.0 mm×10.4 mm) punches. The target mass of the tablet cores was 1550 mg. Samples of the core tablets were taken from a minimum of 20 equally spaced locations during the compression run and analyzed for content uniformity. In general, all powder blends compressed satisfactory on the rotary tablet press with respect to tablet hardness, friability, tablet thickness, tablet appearance, and tablet weight variation. The compression operation was performed at a rate of approximately 500 tablets/minute (12 rpm press speed) or approximately 0.8 kg/minute to deliver satisfactory tablet weight uniformity.

Tablet Film-Coating

Suitable film coatings are selected by routine screening of commercially available preparations. This activity is well within the skill of the ordinary artisan. Each lot of tablet cores was divided into two coating sub-lots that were film coated in a 48-inch Thomas Engineering COMPU-LAB coating pan using a dual-nozzle spraying system. All the tablet cores were film-coated using a 15% w/w aqueous coating suspension Opadry II Pink, which was used within 24 hours of preparation. All tablets cores were coated to a target weight gain of 3.0% using a target spray rate of 180 g/min, which corresponds to a normalized spray rate of 1.5 to 2.3 g/min/kg tablets.

HPLC Assay for Degradation Products

Efavirenz/emtricitabine/tenofovir DF tablets (EFV/FTC/TDF tablets) are assayed by HPLC for EFV, FTC, and TDF using external reference standards. The degradation products of EFV, FTC, and TDF are determined by area normalization with the application of relative response factors, as appropriate. The identity of EFV, FTC, and TDF are confirmed by comparison of their retention times with those of the reference standards.

Standard and Sample Solution Preparation Standard and Sample Solvent 25 mM Phosphate Buffer, pH 3

Weigh and transfer 3.4 g of potassium phosphate monobasic, anhydrous into a 1 L volumetric flask. Add about 800 mL of water and mix until dissolved.

Adjust the pH to 3.0±0.1 with phosphoric acid, then dilute to volume with water.

Sample Solvent (40:30:30: 25 mM Phosphate Buffer, pH 3:Acetonitrile:Methanol)

Combine 400 mL of 25 mM Phosphate Buffer, pH 300 mL of acetonitrile, and 300 mL of methanol and mix. Allow to equilibrate to ambient temperature.

50:50 Acetonitrile:Methanol

Combine 500 mL of acetonitrile and 500 mL of methanol and mix. Allow to equilibrate to ambient temperature.

Standard Solution

Accurately weigh approximately 60 mg of EFV reference standard 20 mg of FTC reference standard, and 30 mg of TDF reference standard and transfer into a 100 mL volumetric flask. Add approximately 80 mL of sample solvent (40:30:30) to the flask and mix or sonicate until dissolved. Dilute to volume with sample solvent (40:30:30) and mix well. The final concentration of each component is approximately 0.6 mg/mL of EFV, 0.2 mg/mL of FTC, and 0.3 mg/mL of 100.

System Suitability Test Solutions Sensitivity Check Standard

Prepare a 10 μg/mL FTU stock solution by accurately weighing out approximately 10 mg of the FTU authentic substance into a 100 mL volumetric flask. Add sample solvent (40:30:30) to approximately 80% of volume and mix or sonicate until dissolved. Dilute to volume with sample solvent (40:30:30) and mix well. Pipet 10 mL of this solution into a 100 mL volumetric flask. Dilute to volume with sample solvent (40:30:30) and mix well.

Prepare the sensitivity check standard containing 0.2 mg/mL of FTC and 0.2 μg/mL of FTU (0.10% relative to FTC). Accurately weigh out 20 mg FTC into a 100 mL volumetric flask. Using a Class A pipet, transfer 2.0 mL of the FTU stock solution into the same flask. Add additional sample solvent (40:30:30) to the flask and mix or sonicate until dissolved. Dilute to volume with sample solvent (40:30:30) and mix well. Alternatively, 2.0 mL of the 10 μg/mL FTU stock solution may be added to the standard solution prior to diluting to volume.

Sample Preparation for EFV/FTC/TDF Tablets

The strength and degradation product content of EFV/FTC/TDF tablets is determined by the analysis of a composite solution prepared from ten tablets.

The final concentration of each component in the sample solution is approximately 0.6 mg/mL of EFV, 0.2 mg/mL of FTC, and 0.3 mg/mL of TDF.

-   -   a) Place ten tablets into a 1 L volumetric flask and add 400 mL         25 mM phosphate buffer, pH 3 to the volumetric flask.     -   b) Mix by stirring vigorously for about 75 minutes.     -   c) Add 50:50 acetonitrile:methanol to the flask to approximately         2 cm below the volume mark.     -   d) Equilibrate the solution to ambient temperature by mixing for         an hour. Dilute to volume with 50:50 acetonitrile:methanol. Mix         well by inverting the flask or stirring with a magnetic stir         bar.     -   e) Using a 0.45 μm syringe filter with a syringe, filter         approximately 10 mL of step (d) for the next dilution. Discard         the first 2 mL of filtrate.     -   f) Using a Class A pipet, transfer 5.0 mL of the filtrate from         step (e) into a 50 mL volumetric flask and dilute to volume with         sample solvent (40:30:30). Mix well.

Chromatography

1. An HPLC equipped with a UV detector and an electronic data acquisition system is used.

2. An HPLC column, 4.6 mm i.d. by 250 mm long, packed with C12 reversed phase, 4 μm particle size, 80 Å pore size material is used.

3. Mobile phase buffer: Prepare a 20 mM ammonium acetate buffer, pH 4.6; adjust pH with acetic acid as needed.

4. Mobile phase gradient: Elute with Mobile Phase Buffer:acetonitrile from 99:1 to 1:99 over 67 minutes.

5. Peak detection: UV at 262 nm

6. Injection volume: 10 μL

Under the stated chromatographic conditions, the retention times of the FTC, TDF and EFV peaks are typically 11, 33, and 50 minutes, respectively.

Injection Sequence

Inject the sample solvent at least twice as a blank to ensure that the column is equilibrated and to identity any potential artifact peaks.

Inject the sensitivity check standard or standard solution containing approximately 0.10% FTU to measure the sensitivity of detection.

Inject five replicates of standard solution 1 (R1), followed by a single injection of standard solution 2 (R2). Calculate the theoretical plates and tailing factors from the standard solution injections.

For identity, strength, and degradation products determination, perform duplicate injections of the sample solution.

All sample solutions must be bracketed by standard solution injections. Generally, not more than ten sample solution injections between bracketing standard injections is recommended.

System Suitability Theoretical Plates and Tailing Factor

Calculate the number of theoretical plates (N) and the tailing factors (T) for the EFV, FTC, and TDF peaks from the Standard Solution Chromatogram. The formulas for N and T determination are defined in the current United States Pharmacopeia. The values of these parameters must conform to the criteria :N≦40,000 and 0.8≦T≧2.0.

Sensitivity Check

The sensitivity check will utilize the FTU peak in the sensitivity check standard present at approximately 0.10%. Calculate the area percent of the FTU peak with the appropriate RRF (listed in Table 2) applied for the sensitivity check standard using the calculation for percent individual degradation product. Compare this result to the theoretical percent of FTU for the sensitivity check standard as follows:

${Sensitivity} = \frac{{FTU}_{Determined}}{{FTU}_{Theoretical}}$

Where

-   -   FTU_(Determined)=area percent of FTU determined for the         sensitivity check standard or standard solution     -   FTU_(Theoretical)=theoretical area percent of FTU for the         sensitivity check standard or standard solution

The sensitivity must be between 0.70-1.30.

Evaluation and Calculations Identification of Degradation Products

Employ the appropriate detection parameters (such as peak threshold, minimum peak area, etc.) to allow detection of peaks present at 0.05% or less. Identify the impurities and degradation products of EFV, FTC, and TDF present in the chromatograms of the sample solution injections by noting the relative retention times (RRT) of the observed secondary peaks, discounting any peaks not related to the sample. Only degradation products are quantified. Calculate the average of the results from all sample solutions injections to the nearest 0.01%. In cases where the degradation product was not detected or was below the threshold of integration in one injection and/or sample, use only the quantified results in the recalculation (i.e., do not treat as a zero value).

${RRT} = \frac{{retention}\mspace{14mu} {time}\mspace{14mu} {of}\mspace{14mu} {the}\mspace{14mu} {secondary}\mspace{14mu} {peak}}{{retention}\mspace{14mu} {time}\mspace{14mu} {of}\mspace{14mu} {the}\mspace{14mu} {tenofovir}\mspace{14mu} {disoproxil}\mspace{14mu} {peak}}$

The RRTs and the relative response factor (RRF) values of the potential impurities and degradation products for EFV are shown in Table 1, and the degradation products are shown in bold-face. The impurities and degradation products for FTC are shown in Table 2, and the degradation products are in bold-face. The impurities and degradation products for TDF are shown in Table 3, and the degradation products are in bold face.

As the RRT may vary, the identity of impurities and degradation products may be confirmed by comparison to authentic substances (or to impurity and degradation product peaks in the reference standard), if required.

Degradation Product Content Determination Quantification of FTC Degradation Products

Determine the level of each degradation product of FTC observed in the chromatograms of the sample solution injections using the following formula:

${{Degradation}\mspace{14mu} {Product}\mspace{14mu} (\%)} = {\frac{1}{TPA} \times {RRF} \times 100}$

Where:

-   -   I=Area of the degradation product peak     -   TPA=Total peak area (area of FTC and all related degradation         products, excluding impurities and artifacts), corrected by RRF.     -   RRF=Relative response factor with respect to FTC

8.4.3 Quantification of TDF Degradation Products

Determine the level of each degradation product of TDF observed in the chromatograms of the sample solution injections using the following formula:

${{Degradation}\mspace{14mu} {Product}\mspace{14mu} (\%)} = {\frac{1}{TPA} \times {RRF} \times 100}$

Where

-   -   I=Area of the degradation product peak or unassigned peak     -   TPA=Total peak area (area of the TDF main peak, all related         degradation products, and all unassigned peaks, excluding         impurities and artifacts), corrected by RRF     -   RRF=Relative response factor with respect to TDF

Results and Reporting Degradation Products Content

Report individually the average of the results for each degradation product observed to the nearest 0.01%. Report the total degradation product content of EFV, FTC, and TDF respectively to the nearest 0.1%, as the sum of the average levels of all degradation product peak observed. For degradation products found at levels less than 0.05%, report their levels as trace and do not include their levels in the calculation of total degradation product content.

REFERENCES

United State Pharmacopeia <621>

Pharmacopeial Forum 26(4) 2000

TABLE 1 EFV related impurities and degradation products EFV Related Approximate Compound RRT^(a) RRF^(b) SD-573 ^(c) 1.46 0.5 SR-695^(d) 1.50 EFV 1.50 SP-234 1.57 SW-965 1.60 SE-563 1.73 SM-097 ^(c) 1.83 0.5 ^(a)Approximate RRTs, and the values are relative to the TDF peak ^(b)RRFs for EFV related degradation products are relative to EFV ^(c)EFV related degradation products ^(d)SR-695 elutes before EFV (approximately 0.1 min separation) Degradation products are marked in bold face

TABLE 2 FTC related degradation product FTC Related Approximate Compound RRT^(a) RRF^(b) FTC 0.33 FTU ^(c) 0.38 0.7 ^(a)Approximate RRTs, and the values are relative to the TDF peak ^(b)RFs for FTC related degradation products are relative to FTC ^(c)FTC related degradation products

TABLE 3 Tenofovir DF related degradation products TDF Related Approximate Compound RRT^(a) RRF^(b) mono-POC PMPA ^(c) 0.47 0.6 Mixed Dimer ^(c) 0.98 1.0 TDF 1.00 Dimer ^(c) 1.34 0.9 ^(a)Approximate RRTs, and the values are relative to the TDF peak ^(b)RRFs for TDF related degradation products are relative to TDF ^(c)TDF related degradation products 

1. A composition comprising tenofovir DF and a surfactant whereby the surfactant is in a stabilizing configuration with the tenofovir DF. 2-30. (canceled) 